Dependence of insulin-stimulated glucose transporter 4 translocation on 3-phosphoinositide-dependent protein kinase-1 and its target threonine-410 in the activation loop of protein kinase C-ζ

G. Bandyopadhyay, M. L. Standaert, M. P. Sajan, Larry M Karnitz, L. Cong, M. J. Quon, R. V. Farese

Research output: Contribution to journalArticle

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Abstract

Previous studies have suggested that 1) atypical protein kinase C (PKC) isoforms are required for insulin stimulation of glucose transport, and 2) 3-phosphoinositide-dependent protein kinase-1 (PDK-1) is required for activation of atypical PKCs. Presently, we evaluated the role of PDK-1, both in the activation of PKC-ζ, and the translocation of epitope-tagged glucose transporter 4 (GLUT4) to the plasma membrane, during insulin action in transiently transfected rat adipocytes. Overexpression of wild-type PDK-1 provoked increases in the activity of cotransfected hemagglutinin (HA)-tagged PKC-ζ and concomitantly enhanced HA-tagged GLUT4 translocation. Expression of both kinase-inactive PDK-1 and an activation-resistant form of PKC-ζ that is mutated at Thr-410, the immediate target of PDK-1 in the activation loop of PKC-ζ, inhibited insulin-induced increases in both HA-PKC-ζ activity and HA-GLUT4 translocation to the same extent as kinase-inactive PKC-ζ. Moreover, the inhibitory effects of kinase-inactive PDK-1 were fully reversed by cotransfection of wild-type PDK-1 and partly reversed by wild-type PKC-ζ, but not by wild-type PKB. In contrast to the T410A PKC-ζ mutant, an analogous double mutant of PKB (T308A/S473A) that is resistant to PDK-1 activation had only a small effect on insulin-stimulated HA-GLUT4 translocation and did not inhibit HA-GLUT4 translocation induced by overexpression of wild-type PDK-1. Our findings suggest that both PDK-1 and its downstream target, Thr-410 in the activation loop of PKC-ζ, are required for insulin-stimulated glucose transport.

Original languageEnglish (US)
Pages (from-to)1766-1772
Number of pages7
JournalMolecular Endocrinology
Volume13
Issue number10
StatePublished - 1999

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3-Phosphoinositide-Dependent Protein Kinases
Facilitative Glucose Transport Proteins
Threonine
Protein Kinases
Protein Kinase C
Phosphatidylinositols
Hemagglutinins
Insulin
1-Phosphatidylinositol 4-Kinase
Glucose
Adipocytes
Epitopes
Protein Isoforms

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology, Diabetes and Metabolism

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Dependence of insulin-stimulated glucose transporter 4 translocation on 3-phosphoinositide-dependent protein kinase-1 and its target threonine-410 in the activation loop of protein kinase C-ζ. / Bandyopadhyay, G.; Standaert, M. L.; Sajan, M. P.; Karnitz, Larry M; Cong, L.; Quon, M. J.; Farese, R. V.

In: Molecular Endocrinology, Vol. 13, No. 10, 1999, p. 1766-1772.

Research output: Contribution to journalArticle

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abstract = "Previous studies have suggested that 1) atypical protein kinase C (PKC) isoforms are required for insulin stimulation of glucose transport, and 2) 3-phosphoinositide-dependent protein kinase-1 (PDK-1) is required for activation of atypical PKCs. Presently, we evaluated the role of PDK-1, both in the activation of PKC-ζ, and the translocation of epitope-tagged glucose transporter 4 (GLUT4) to the plasma membrane, during insulin action in transiently transfected rat adipocytes. Overexpression of wild-type PDK-1 provoked increases in the activity of cotransfected hemagglutinin (HA)-tagged PKC-ζ and concomitantly enhanced HA-tagged GLUT4 translocation. Expression of both kinase-inactive PDK-1 and an activation-resistant form of PKC-ζ that is mutated at Thr-410, the immediate target of PDK-1 in the activation loop of PKC-ζ, inhibited insulin-induced increases in both HA-PKC-ζ activity and HA-GLUT4 translocation to the same extent as kinase-inactive PKC-ζ. Moreover, the inhibitory effects of kinase-inactive PDK-1 were fully reversed by cotransfection of wild-type PDK-1 and partly reversed by wild-type PKC-ζ, but not by wild-type PKB. In contrast to the T410A PKC-ζ mutant, an analogous double mutant of PKB (T308A/S473A) that is resistant to PDK-1 activation had only a small effect on insulin-stimulated HA-GLUT4 translocation and did not inhibit HA-GLUT4 translocation induced by overexpression of wild-type PDK-1. Our findings suggest that both PDK-1 and its downstream target, Thr-410 in the activation loop of PKC-ζ, are required for insulin-stimulated glucose transport.",
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